New genetic links in eosinophilic esophagitis

Eosinophilic esophagitis (EoE) is increasingly diagnosed as a disorder throughout the world. It is characterized by eosinophils in the esophagus due to food allergies. Molecular analysis of esophageal biopsies and mouse models have indicated a clear role for the T helper 2 pathway, in particular interleukins 5 and 13, in this disease. Current treatment options for EoE involve avoidance of the allergens or using anti-inflammatory medications such as topical corticosteroids. In the past year, genomic research has led to the identification of single nucleotide polymorphisms in the gene encoding thymic stromal lymphopoietin (TSLP), and subsequently in the gene encoding its receptor, as disease susceptibility markers for EoE. Identification of this molecule and its receptor suggest the potential for new treatment options in the future.


Definition and prevalence of eosinophilic esophagitis
Eosinophilic gastrointestinal diseases are increasingly recognized and diagnosed disorders that include eosino philic esophagitis (EoE), eosinophilic gastroenteritis and eosinophilic colitis. Among the eosinophilic gastro intes tinal diseases, EoE has become increasingly prevalent in the United States, Switzerland and Australia [1]. Overall, EoE is a global health condition now reported on all continents except Africa, with an incidence of approximately 5 in 10,000 [1,2].
The diagnosis of EoE is based on clinicopathology and depends in part on demonstration of esophageal eosino philia. A panel of experts proposed consensus recom men dations in 2007 [1], which included the demon stration of 15 or more eosinophils in at least one highpowered field despite treatment with a proton pump inhibitor or exclusion of gastroesophageal reflux disease (GERD) by ambulatory pH monitoring. The clinical presentation of EoE varies with age: infants and toddlers often have difficulty feeding and fail to thrive, schoolage children present with vomiting and epigastric or chest pain, and adolescents and adults present with dysphagia and food impaction [3].
EoE is considered a foodallergyrelated disorder on the basis of several findings. Most patients are atopic individuals (that is, they have a high rate of food allergen sensitization as determined using skin prick and patch testing); in addition, EoE patients have a higher rate of food anaphylaxis than the general population [4]. More over, nearly all EoE patients have complete remission following introduction of an elemental formula diet that removes all allergens from the diet; conversely, the disease flares on reintroduction of specific foods [3]. Unlike classic anaphylaxis that typically involves a limited set of foods, EoE patients are often sensitized to a myriad of foods, often including food groups not typically considered to elicit anaphylaxis [4].

Mechanism of eosinophilic esophagitis
EoE seems to be similar to many other allergic diseases with a mechanism of disease mediated by T helper 2 (Th2) cells. Experimental mouse models have demon strated key roles for adaptive immunity, Th2 cell cyto kines (especially interleukin (IL)5 and IL13) and eosinophilicattraction chemokines such as CCL26 (also called eotaxin3) in the development of EoE, and also a strong connection between allergic sensitization and inflammation in the respiratory tract and skin [5,6]. IL13 has been shown to induce many of the features of EoE in human tissue and murine systems, including the induc tion of CCL26 [7]. Genomewide profiling revealed that CCL26 is overexpressed about 50fold compared with normal controls or patients with GERD [6]. In addition, Rothenberg and colleagues [6] found that the EoE transcriptome was consistent across sex, age and familial or nonfamilial inheritance patterns and was independent of atopic status, suggesting a common disease mechanism despite phenotypic variations. A potential proposed mechanism is that Th2 cell activation leads to over expression of CCL26 and thereby to migration of eosinophils to the esophagus.
Work by Aceves and colleagues [8] showed that eosinophilic migration and activation in EoE lead to

Abstract
Eosinophilic esophagitis (EoE) is increasingly diagnosed as a disorder throughout the world. It is characterized by eosinophils in the esophagus due to food allergies. Molecular analysis of esophageal biopsies and mouse models have indicated a clear role for the T helper 2 pathway, in particular interleukins 5 and 13, in this disease. Current treatment options for EoE involve avoidance of the allergens or using anti-inflammatory medications such as topical corticosteroids. In the past year, genomic research has led to the identification of single nucleotide polymorphisms in the gene encoding thymic stromal lymphopoietin (TSLP), and subsequently in the gene encoding its receptor, as disease susceptibility markers for EoE. Identification of this molecule and its receptor suggest the potential for new treatment options in the future.
subepithelial fibrosis and to increased expression of transforming growth factor β (TGFβ) and its down stream signaling molecule phosphoSMAD2/3 compared with patients with GERD and healthy controls. In addition, esophageal biopsies demonstrated an increased expression of vascular cell adhesion molecule 1 (VCAM1) [8]. This fibrosis may account for the esophageal dysmotility that leads to the symptoms of dysphagia and food impaction observed in adults. The chronic fibrosis and changes in inflammatory markers also parallel symptoms observed in asthma.

Genetic links in EoE
The evidence indicates that EoE has a strong familial association, with nearly 10% of parents of EoE patients having a history of esophageal strictures and about 8% having biopsyproven EoE [5]. EoE also shows a high sibling risk ratio (λ S ) of approximately 80 compared with related atopic diseases such as asthma (λ S about 2) [9]. The first candidate gene for EoE identified was CCL-26, the most overexpressed gene in the esophagus as determined by genomewide expression profiling [6]. However, the diseaseassociated allele is present in only 14% of EoE patients [6].
Rather than looking at the potential pathways, Aceves and colleagues [10] examined response to topical corticosteroids a standard therapy for the condition in patients with EoE. A positive response was defined as residual eosinophil counts of seven or fewer eosinophils per highpower field. Responders had a reduced esophageal remodeling with decreased fibrosis, fewer TGFβ and pSmad2/3positive cells and decreased vascular activation following therapy with the cortico steroid budesonide. Responders were more likely to have a CC genotype at the 509 position in the TGFβ promoter than were nonresponders.
More recently, a multicenter genomewide association study (GWAS) has identified the thymic stromal lympho poietin (TSLP) gene, at 5q22, as an important candidate gene in the pathogenesis of the disease [11]. TSLP is an epithelialderived cytokine that activates professional antigenpresenting cells, such as dendritic cells, which initiate Th2type allergic responses [12]. The same study also found an increased expression of TSLP in the esophagi of patients with EoE compared with healthy controls [11].
The most recent finding by Sherrill et al. [13] adds to our understanding of the interaction of TSLP and EoE. This genomic analysis [13] focused on key allergic single nucleotide polymorphisms (SNPs) along with epithelial SNPs and compared them in atopic and healthy controls. In addition to replicating the significant association between SNPs at TSLP and EoE, they found that TSLP variants remained significant when compared with atopic controls in their study [13] or asthma controls from the previous GWAS analysis [11]. However, variations in the TSLP gene were not significant when compared with controls, probably because of the small number of controls used. TSLP has also been found to be associated with atopic dermatitis [14] and asthma [15] compared with healthy controls. The finding of TSLP being significant only among the atopic controls could be due either to the other studies not removing patients with EoE (which is commonly atopic) from their control group or to the possibility that TSLP might trigger multiple atopic diseases depending on secondary signals.
Sherrill and colleagues [13] also found a SNP in the TSLP receptor (TSLPR) gene that was associated with EoE. The gene encoding the TSLP receptor is located on the pseudoautosomal region on Xp22.3 and Yp11.3 and it was significantly associated with disease only in male patients with EoE. Given that EoE is more common among males by a 2:1 ratio [1], the finding of a SNP in the TSLPR suggests a potential mechanism for the male dominance observed in EoE. Sherrill et al. [13] also noted one additional finding that may suggest alternative pathways for the treatment or the pathogenesis of EoE: they observed that stimulation of primary esophageal epithelial cells with poly I:C (a doublestranded RNA mimetic) induced the expression of TSLP mRNA. This induction was dependent on Tolllike receptor (TLR)3 stimulation. TLR3 recognizes doublestranded RNA, which is found in some viruses such as reoviruses. These results suggest a second hit for the development of EoE with a viral trigger and allergen exposure. In the lung and skin TSLP is produced primarily by epithelial cells in response to Th2 cytokines or TLR3 agonists, and it subsequently targets dendritic cells to secrete Th2 inducing activity, including Th2 cytokine and chemokine production.
These findings [13] suggest a unique potential mecha nism for the induction of EoE. Could food allergens trigger the TLR3 receptor, inducing TSLP and causing the activation of the Th2 pathway, leading to eosinophilic inflammation in the esophagus? Or is there a 'second hit' , a virus that makes susceptible people develop EoE?

Potential pathways for treatment
The current treatment options for EoE involve the avoidance of the trigger (foods) and the treatment of the underlying inflammation with topical corticosteroids [1]. Treatment with food avoidance is highly successful, with rates close to 100% with elemental diets (amino acid formulas) [1]. However, these formulas are unpalatable and lead to low quality of life. Eliminating foods on the basis of allergy testing or empirical elimination leads to resolution of esophageal eosinophilia in 50 to 80% of patients, depending on the diet and the age of the patient and reduction of symptoms in over 90% of the patients [3]. These diets are also difficult to maintain and many patients refuse to continue them. Treatment with topical steroids can work for 50 to 80% of patients, but there are also some drawbacks. Topical corticosteroids can lead to localized yeast infections and have potential longterm side effects, including growth suppression and osteopenia (low bone density); however, these have not been studied or seen in shortterm studies. Other treatments currently being investigated include antiimmunoglobulin (Ig)E, antiIL5, antiIL13 and chemoattractant homologous receptor expressed on Th2 cells (CRTH2) antagonist or topical corticosteroids [1618].
The recent identification of TSLP and its receptor as key components in the EoE pathogenesis [13] suggests that blockage of the TSLPTSLP receptor activation could provide an attractive approach to treating the cause of EoE. In addition, the new finding that TLR3 induces TSLP suggests that a second hit (a virus) may trigger this pathway. If a particular virus or microorganism unique for EoE were identified, this would allow the development of a preventive strategy. This is unlikely to occur in the near future, but treatment with TLR3 antagonists or blocking downstream signaling already represent hope for the treatment of EoE.